1. Field of the Invention
This invention lies in the field of the growth and functionality of pancreatic cells.
2. Description of the Prior Art
Type I (insulin-dependent) diabetes mellitus is a widespread disease, arising from an autoimmune disorder in which insulin-secreting beta-cells in the pancreas are destroyed. The loss of these cells impairs the body's ability to assimilate glucose from the blood, and the resulting high glucose levels can lead to blindness, kidney disease, nerve damage, and ultimately death. Insulin injections are commonly used to compensate for the lack of beta cells, but blood sugar levels can still fluctuate widely. Methods of lessening the fluctuations have included the use of small, frequent doses of insulin and the use of mechanical pumps that mimic the action of the pancreas, but these require continuous or periodic maintenance, and the results are often of limited success. An alternative is a pancreatic transplant, but this requires major surgery and the availability of donor pancreases is limited.
A more promising option is the transplantation of islets of Langerhans, using tissue derived from either cadavers or human fetuses. Islets of Langerhans are clusters of cells in the pancreas that include the insulin-secreting beta cells, and their transplantation entails considerably less risk than the transplantation of a pancreas. Sources for islet transplantation include adult pancreatic tissue, fetal pancreatic tissue and islet-like cell clusters (ICCs). Fetal tissue offers a greater content of islets in proportion to its mass, as well as a greater capacity for proliferation with its less mature cells. Islet-like cell clusters are heterogeneous cell populations that include epithelial cells that differentiate after transplantation to form various types of cells including mature islets.
Islet tissue that is available for transplantation is scarce, however, and islets must be banked and transported in order to obtain sufficient islets for a single recipient. One means of permitting the accumulation needed to obtain a sufficient number of islets is cryopreservation. Ex vivo or in vitro proliferation, or the growth of islet tissue in culture media, is another. Ex vivo or in vitro proliferation are also of use in generating sufficient islet tissue for clinical and laboratory research.
Successful ex vivo or in vitro proliferation occurs when the proliferated cells retain their functionality as sources of insulin. Regulation of the proliferation and differentiation of human fetal pancreatic islet cells is dependent on interactions between cell-cell and cell-matrix contacts and specific growth factors. Unfortunately, the nature of these interactions, and in particular the relative effects of matrix and growth factors on growth vs. differentiation, are not known. This prevents one from selecting matrices and growth factors, or combinations of the two, that will achieve proliferation while retaining cell functionality.
It is known, for example, that cell division among adult human .beta.-cells is induced by a combination of extracellular matrix and hepatocyte growth factor/scatter factor (HGF/SF), the latter being one of a variety of peptide growth factors that have been tested. The cell proliferation that is achieved, however, is accompanied by a downregulation of islet-specific gene expression. Cell transplantation experiments performed with a cell suspension of proliferated cells failed to produce mature endocrine cells. The use of nicotinamide, a potent inducer of endocrine differentiation, in a suspension of the cells resulted in new islet formation, but this was accompanied by considerable cell attrition, which canceled much of the benefit of the new cell formation.
A method and growth medium are therefore sought that will result in the proliferation of islets and ICCs at a substantial rate with minimal or no loss of insulin secreting function.